A Project to Determine the Best Option for the Long-Term Self-Sustainability of the Canadian Swine Health Intelligence Network (CSHIN)

Graham Clarke11Canadian Pork Council

This project is one of four projects governed under the terms of a Contribution Agreement between the Canadian Swine Health Board (CSHB) and the Minister of Agriculture and Agri-Food that, when combined, have the overall objective to develop, implement, deliver and maintain a framework that would lead to the establishment of long-term disease management solutions for the hog industry.

This particular project is to evaluate two specific options that will provide for the long-term self-sustainability of the Canadian Swine Health Intelligence Network (CSHIN). The specific options to be evaluated are:

1. Continuing the activities of the CSHIN under the CSHB 2. Moving the CSHIN under the Canadian Animal Health Surveillance Network [CAHSN]

Any other potential options that may be identified during the process will also be evaluated.

Consultants from Serecon Management Consulting Inc. have been engaged to conduct the work and it is expected that a recommendation will be made at the end of June, 2014.

CAHSN & Enabling National Surveillance Coordination

Christine Power11Animal Health Science Directorate, CFIA

Over the past fifteen years, the Canadian landscape has been enriched by significant multi-organisational collaborations in the domain of animal health such as: CAHLN(2002);CAHC(2003);CCVO(2005);CAHSN(2005);NFAHWC(2009)

Each organisation was created to meet a pressing need that encompassed challenges beyond the existing capacity of any one organisation to resolve on its own. Over the years, not without with great effort by its founders and leaders, these organisations have tackled the bigger-than-life issues and charted a course to mutual understanding, team work and creatively building joint solutions. The synergies, opportunities and challenges of official collaborations have become very familiar to many of us, such that, for many, it is the way we do business day to day.

While obtaining current information on the health and disease status of Canadian livestock and poultry is the aspiration of a multitude of animal health stakeholders, it has been difficult to achieve. Several reports commissioned by the CCVO and NFAHWC in recent years indicate that the absence of national coordination for intelligence, monitoring and surveillance activities is a major stumbling block. Since governance is key for any new structure, the NFAHWC is currently leading a multi-stakeholder process to clarify the roles, responsibilities and location of a national surveillance coordination group. Close attention to governance design at the outset is an important first step towards achieving the collective goal of timely access to national animal disease surveillance information.

Methicillin-resistant Staphylococcus pseudintermedius (MRSP) has emerged as a major pathogen in dogs. The detection of MRSP in clinical specimens is critical in improving and maintaining patient health. The objective of this study was to determine if using selective culture methods would detect more MRSP compared to traditional culture methods. Samples were submitted to the AVC Diagnostic Bacteriology Laboratory for routine culture and susceptibility testing. Dogs considered high-risk for MRSP infection were included in the study. Samples were cultured using four methods: traditional agars, mannitol salt agar with 2µg/mL oxacillin (MSAox), mannitol salt enrichment broth (EB) with MSAox, and EB with traditional agars. MRSP was identified using conventional methodology, and MRSP was confirmed using identical procedures regardless of culture method. Prevalence of MRSP was estimated for each test method, and discordance between test methods was evaluated. Analysis based on 752 samples collected between February 2013 and March 2014 estimated the prevalence of MRSP if any test method was positive at 13.3%. In the same data set, the prevalence of MRSP was estimated by traditional agars at 10.1%, by MSAox at 8.9%, by EB and traditional agars at 10.8%, and by EB and MSAox at 11.2%. No statistically significant differences in MRSP recovery were found between traditional agars and MSA (p=0.208) or EB and MSAox (p=0.223) or with EB (p=0.322). Further investigation of MSAox with and without EB as a rapid diagnostic test is in progress.

Detection of Mg and Ms DNA in samples submitted to The Georgia Poultry Lab using IDEXX Real Time PCR reagents

Kristen Roza11Livestock, Poultry and Dairy Division, IDEXX

Detection of DNA from pathogenic Avian Mycoplasmas can be challenging due to the sophisticated techniques required to achieve accurate results. Inhibition of the reaction as a result of sample contamination can occur at any step throughout the process. Most real time PCR (qPCR) assays fail to identify if inhibitors have affected the reaction and positive results may be errantly classified as negative. The Georgia Poultry Lab uses commercially available IDEXX real time PCR reagents to detect Mg and Ms DNA. These reagent sets contain an internal control, meant to detect the presence of inhibitors in the reactions. In this poster, results are reviewed. The use of this qPCR format involving hybridization probes and an internal control in each individual well is evaluated and discussed.

Diagnostic challenge for the detection of Campylobacter fetus subspecies venerealis in cattle

The genus Campylobacter contains multiple species including two subspecies of C. fetus: C. fetus ssp. fetus (CFF) and C. fetus ssp. venerealis (CFV). Culture and differentiation of CFF and CFV remain difficult due to the fastidious growth requirements and biochemical inactivity of these organisms. CFV has adapted to the genital tract of bulls and causes bovine genital campylobacteriosis in cows. The disease is notifiable by the Office International des Epizootie (OIE), because of its negative economic impact; therefore, animals must be certified CFV-free for international trade. Bulls are asymptomatic carriers; however, CFV infection in cows leads to reproductive failure due to early embryonic death, abortion or infertility. Detection of carrier bulls is very important for disease control in a herd. It is recommended to culture samples [preputial washes (PWs)] within 4 hrs after collection as CFV is very fragile outside the host. In case of delays during shipment to the lab, PWs can be inoculated into transport enrichment medium (TEM) prior to culture.For the past few years, we have used various TEM for sample collection, shipment to the lab and further culture, but with mixed success.In the present study, we evaluated the recovery rate of CFV using Weybridge and Landers TEM, followed by culture on blood agar and Skirrow’s media. Plates were incubated under micro-aerophilic condition (6% O2, 10% CO2 and 84% N2), for up to 5 days and examined for bacterial growth. Culture recovery rate was compared to the detection of CFV using a SYBR Green real time PCR (rt-PCR) assay performed on PWs. These were collected in Phosphate Buffered Saline (PBS) or “InPouch TF“ bovine medium used for Tritrichomonas foetus testing, incubated at 35°C for 48 hrs.The recovery rate of CFV, using TEM and culture, was low (0.6%), compared to the detection by rt-PCR from PWs submitted in PBS or “InPouch TF” (8.0%). These results show that rt-PCR test may be considered for CFV detection.

DNA sexing and phylogeny studies in birds of prey and parrots in Quebec

Knowing the sex of birds is important for diverse biological disciplines including veterinary, ecology and studies of evolution. For bird breeders, knowing the sex of their birds is primordial in order to form reproductive breeding couples. For many species of birds, including juveniles and adults, there is little or no external dimorphism between the sexes. Numerous techniques can be used to sex birds, but each technique has its own disadvantages, which may include invasive manipulation of the animal. Using blood or tissue samples is a rapid method to determine the sex of birds via DNA analysis. Unfortunately, the current reliability of these tests can be questioned.

The objective of the current study is to accurately identify the sex of birds of prey in Quebec, and of parrots seen in veterinary clinics, using DNA sequences of the CHD (chromobox helicase DNA binding) gene, which is found on the avail sex chromosomes W and Z. DNA was extracted from a range of parrot and bird of prey species. Intron 9-10 of the CHD nuclear gene was amplified by PCR. By sequencing the resulting amplicon, the sex of the animal can be easily determined: a single sequence representing CHD-Z indicates a male bird, while a double sequence (representing overlapping CHD-Z and CHD-W sequences) indicates a female bird. Distinct CHD-W sequences were obtained by sub-cloning female amplicons into bacterial vectors. The DNA sequences obtained for the CHD-Z and CHD-W genes were deposited in public domain DNA repositories. In parallel to DNA sexing based on the CHD nuclear gene, the species of the bird was verified based on the COX1 mitochondrial gene. Using COX1, CHD-Z and CHD-W sequences, phylogenetic studies can be performed for the bird species tested.

DSP - Disease Surveillance Program – the Ontario plan

Animal health and public health depend upon a high level of awareness of the ever-changing status of endemic, new, and emerging diseases. In several current systems, such as RAIZO and CSHIN, passively acquired laboratory data is combined with clinical impressions from veterinary practitioners to generate a clear picture of current conditions in the various commodities. In a new 5-year plan, 2013-2018, the AHL has been funded by OMAF/MRA to fulfil 4 objectives: provide an environmental scan of the current state of animal health surveillance, enhance Ontario disease surveillance, integrate with national surveillance, and improve laboratory testing tools. To date, we have developed a framework for a RAIZO-type plan in Ontario, in which species-specific “expert networks” comprising private practice government, lab, and OVC veterinarians collaborate in providing timely information to stakeholders. We are also working with partners in CFIA and across Canada to develop demonstration examples of integrating provincial disease information into a national database. This proposed network approach builds on the base of early detection and extension related to important animal health hazards in Ontario.

Enteric Zoonoses and Pet Food: A Public Health Survey

Erin Leonard11Public Health Agency of Canada

Background: Contact with animals, their food, and their environment is an important source of human illness for pathogens such as Campylobacter, VTEC, and Salmonella. Recent outbreaks of disease associated with exposure to pets and pet foods have highlighted some challenges inherent in pet-related enteric outbreak investigations. A better understanding of current legislation, practices, and challenges could guide future recommendations to enhance the efficiency of public investigations and resultant public health interventions. Methods: To get a better understanding of current practices and the challenges associated with pet-related outbreaks across Canada, a survey was conducted in the fall of 2013. Interviews were conducted with provincial and territorial public health officials; questions pertained to legislation, data collection, outbreak investigation, collaboration, significance, and prevention and control. Key findings: Pets and pet foods/products were ranked relatively low in significance relative to foodborne as the source of enteric illness, however all jurisdictions recognized that pet-related outbreaks are preventable; collaboration is a regular occurrence in pet-related outbreaks however the lack of clarity of roles and responsibilities hinders investigations and poses significant challenges across Canada; there is no systematic collection of animal-type exposures during routine case follow up; although most jurisdictions could identify a lab to which pet-related samples could be sent, the process for doing so is unclear in many jurisdictions. Conclusions: Engagement of public health and animal health partners across Canada in clarifying roles and responsibilities through the development of an outbreak response protocol is critical to improve outbreak detection and lead to a better understanding of the burden of disease related to exposure to pets, pet food and pet products in Canada.

Evaluation of seven paratuberculosis diagnostic tests in the dairy small ruminant industries of Ontario.

A cross-sectional study of the Ontario dairy sheep and dairy goat populations was performed between October 2010 and August 2011. The objective was to estimate the test sensitivity and specificity of seven commercially available paratuberculosis tests: faecal culture, faecal polymerase chain reaction (PCR), two serum enzyme-linked immunosorbent assays (ELISAs), agar gel immunodiffusion (AGID) and two milk ELISAs.

Twenty-nine dairy goat herds were selected using stratified random sampling based on herd size, and 21 dairy sheep flocks were conveniently selected. Twenty lactating animals over the age of two years were randomly selected from each farm resulting in 580 samples from lactating does and 397 samples from lactating ewes. Each animal was sampled for faeces, blood, and milk.

Statistical analysis was performed using frequentist methods and also using latent class analysis (LCA) with Bayesian methodology. Faecal culture had the highest sensitivity in the goat population (81.1%), while in the sheep population, faecal culture demonstrated a sensitivity of 49.5% and faecal PCR a sensitivity of 42.4%. Faecal culture in both populations had an estimated specificity of 98.1% in goats and 97.4% in sheep while the specificity for PCR in both species was <90.0%. Both serum ELISAs in sheep and goats demonstrated higher sensitivities than the serum AGID.

Hepatic alveolar echinococcosis in a dog in Ontario, Canada

A 2-year old neutered male Boxer dog presented to a small animal clinic in southern Ontario with a 12-hour history of anorexia and lethargy. Laparotomy revealed a hemorrhagic, destructive, liver mass. Histopathology was suggestive for an intermediate stage of a cestode. Molecular testing was performed on DNA extracted from formalin-fixed tissue. Sequencing of PCR-generated fragment of the mitochondrial 12S rRNA gene, and RFLP analysis of PCR-generated fragments of mitochondrial 12S rRNA and NADH dehydrogenase 1 genes confirmed the diagnosis of hepatic metacestode of Echinococcus multilocularis. Hepatic alveolar echinococcosis (HAE) in dogs is an extremely rare diagnosis in North America. One case has previously been reported in British Columbia. This is the first reported case in Ontario. It is important that veterinarians recognize that dogs can act as intermediate and definitive hosts for E. multilocularis, and that this tapeworm poses a serious zoonotic threat.

After receiving notification from the NVSL, Ames, IA of the presence of porcine epidemic diarrhea (PED) in the USA, NCFAD, Winnipeg began to prepare for its incursion into Canada. In addition to exchanging PCR protocols, initial preparations also involved obtaining virus, antisera and monoclonal antibodies that could be used in the development of serodiagnostic assays. PEDV propagated in Vero cells together with monoclonal antibodies specific for the PEDV spike glycoprotein obtained from the Central Veterinary Institute Wageningen UR, the Netherlands were used to develop a complex-trapping-blocking ELISA. In January 2014, approximately nine months following the initial detection of PEDV in an Ohio swine herd, the first case of PED was diagnosed in a swine herd in southwestern Ontario. A follow up epidemiological investigation carried out on the Ontario PED cases pointed to feed as a common risk factor. As a result, several lots of feed and spray dried porcine plasma, used as a feed supplement, were tested for the presence of PEDV by real-time RT-PCR assay. Several of these tested positive supporting the notion that contaminated feed may have been responsible for the introduction of PEDV into Canada. These findings led us to conduct a bioassay experiment in which three PEDV positive plasma and two PEDV positive feed samples were used to orally inoculate 3-week-old piglets. Although the feed inoculated piglets did not show any significant excretion of PEDV, the plasma inoculated piglets shed PEDV at a relatively high level for ≥ 9 days. PEDV positive PCR results were confirmed by immunohistochemistry and seroconversion. Despite the fact that PEDV contaminated feed did not result in piglet infection in our bioassay experiment, feed cannot be ruled out as a potential source of infection in the field where other variables may play a contributing role.

PED: A case study of one lab's response to an emerging disease

One of the primary functions of an animal health laboratory is to detect emerging diseases quickly. In the case of porcine epidemic diarrhea virus (PEDV), the rapid spread of PEDV in the US during 2013 was a warning that an incursion of PEDV into Canada was likely just a matter of time. Hence, in August, 2013, the AHL put in place a triplex porcine coronavirus PCR test to detect and differentiate PEDV from TGEV and PRCV, both of which are endemic in Ontario. The index case of PED in Ontario was presented on January 22, 2014 – 4 diarrheic suckling piglets from a 500 sow farrow-to-finish operation submitted to the AHL tested positive for PEDV. Positive samples were also rushed to NCFAD, Winnipeg for additional testing and were confirmed as PEDV-positive on Jan 23. As of early May, 2014, 58 sites are positive in Ontario.

Subsequent to the discovery of porcine deltacoronavirus (PDCoV) in the US in February, 2014, PDCoV was detected by PCR in Ontario in March, 2014 – 6 sites are confirmed positive.

Excellent support and collaboration among AHL, American and CFIA laboratories has allowed early detection of PEDV and PDCoV, and hence an early response to limit the impact of these emerging pathogens.

Provincially led National CAHSN Surveillance Projects

Harold Kloeze11Animal Health Science, CFIA

The Canadian Animal Health Surveillance Network (CAHSN) is a national network of public sector animal health laboratories in Canada that was formed in 2005 in order to provide:1. An early warning surveillance system for animal disease threats to animal health, human health and the security of the food supply;2. An integrated federal-provincial-territorial laboratory network for the diagnosis of serious infectious diseases of animals; 3. A common information-sharing platform for linking federal and provincial animal health laboratories, agencies and departments. CAHSN utilizes the Canadian Network for Public Health Intelligence (CNPHI) platform for this purpose.

A project was designed to collect national data using the CNPHI/CAHSN information platform for three important endemic diseases in each of three species/production groups. The goals of the project were to increase the use of the platform, to solve technical issues with respect to data transmission, to compare the data sets available from the various laboratory partners in the network, to begin the process of report generation and to make recommendations on future direction with respect to each of these topics.

The project was split into three sub-projects based on the species/production groups. British Columbia successfully applied for Growing Forward 2 funding for a project focussing on poultry, Saskatchewan for beef cattle and Ontario (as part of a larger surveillance initiative) for swine. In each case the projects will advance surveillance in the province, improve data transfer to the national platform and act as a nexus for national surveillance for three important endemic diseases in the selected species. National coordination of the projects on a technical level has been instituted in order to provide technical support, provide a forum for collaboration between the sub-projects and to maintain the goals of the originally envisioned national project as described above.

Unusual microbial isolates from the urine of dogs with urinary tract disease

James Goltz11Agriculture, Aquaculture and Fisheries, New Brunswick Provincial Veterinary Laboratory

Disease problems affecting the urinary tract are very common in domestic animals, especially companion animals. Urinalysis and microbial cultures are among the most common requests for laboratory diagnostic testing in specimens from companion animals, and veterinarians and owners are eager for these results to be significant so problems can quickly resolved with the least possible invasiveness, cost, time, inconvenience and animal welfare impact. Interpreting the significance of microbial culture results from urine samples can be challenging, since many of the most significant pathogens are organisms commonly found in the gut or environment. Interesting and unusual microbial culture results from two submissions to the Provincial Veterinary Laboratory will be presented and discussed, including isolates of Listeria monocytogenes and Mycoplasma spumans from an 8 year old Doberman dog with chronic hematuria, and Salmonella infantis and Staphylococcus intermedius group from a 10 year-old Maltese dog with urolithiasis.